Despite advances in treatment, congestive heart failure remains an important cause of mortality in Western countries. Heart failure affects 5 million individuals in the United States alone, and is characterized by a 5-year mortality rate of ˜50% (Levy et al., Long-term trends in the incidence of and survival with heart failure. N. Engl. J. Med., 347:1397-402, 2002).
An important hallmark of heart failure is reduced myocardial contractility (Gwathmey et al., Abnormal intracellular calcium handling in myocardium from patients with end-stage heart failure. Circ. Res., 61:70-76, 1987). In healthy heart muscle, and in other striated muscle, calcium-release channels on the sarcoplasmic reticulum (SR), including ryanodine receptors (RyRs), facilitate coupling of an action potential to a muscle cell's contraction (i.e., excitation-contraction (EC) coupling). Contraction is initiated when calcium (Ca2+) is released from the SR into the surrounding cytoplasm. In heart failure, contractile abnormalities result, in part, from alterations in the signaling cascade that allows the cardiac action potential (AP) to trigger contraction. In particular, in failing hearts, the amplitude of the whole-cell Ca2+ transient is decreased (Beuckelmann et al., Intracellular calcium handling in isolated ventricular myocytes from patients with terminal heart failure. Circ., 85:1046-55, 1992; Gomez et al., Defective excitation-contraction coupling in experimental cardiac hypertrophy and heart failure. Science, 276:800-06, 1997), and the duration is prolonged (Beuckelmann et al., Intracellular calcium handling in isolated ventricular myocytes from patients with terminal heart failure. Circ., 85:1046-55, 1992).
Cardiac arrhythmia, a common feature of heart failure, is known to be associated with SR Ca2+ leaks in structurally-normal hearts. In these cases, the most common mechanism for induction and maintenance of ventricular tachycardia is abnormal automaticity. One form of abnormal automaticity, known as triggered arrhythmia, is associated with aberrant release of SR Ca2+, which initiates delayed after-depolarizations, or DADs (Fozzard, H. A., After depolarizations and triggered activity. Basic Res. Cardiol., 87:105-13, 1992; Wit and Rosen, Pathophysiologic mechanisms of cardiac arrhythmias. Am. Heart J., 106:798-811, 1983). DADs are abnormal depolarizations in cardiomyocytes that occur after repolarization of a cardiac action potential. The molecular basis for the abnormal SR Ca2+ release that results in DADs has not been fully elucidated. However, DADs are known to be blocked by ryanodine, providing evidence that RyR2 may play a key role in the pathogenesis of this aberrant Ca2+ release (Marban et al., Mechanisms of arrhythmogenic delayed and early afterdepolarizations in ferret ventricular muscle. J. Clin. Invest., 78:1185-92, 1986; Song and Belardinelli, ATP promotes development of afterdepolarizations and triggered activity in cardiac myocytes. Am. J. Physiol., 267:H2005-11, 1994).
The most common cardiac arrhythmia in humans is atrial fibrillation (AF). It represents a major cause of morbidity and mortality (Chugh et al., Epidemiology and natural history of atrial fibrillation: clinical implications. J. Am. Coll. Cardiol., 37:371-78, 2001; Falk, R. H., Atrial fibrillation. N. Engl. J. Med., 344:1067-78, 2001). Despite its clinical importance, however, treatment options for AF have been limited—due, in part, to the fact that its underlying molecular mechanisms are poorly understood.
Approximately 50% of all patients with heart disease die from fatal cardiac arrhythmias. Such fatal cardiac arrhythmias are often ventricular in nature. In some cases, a ventricular arrhythmia in the heart may be rapidly fatal—a phenomenon referred to as “sudden cardiac death” (SCD). Fatal ventricular arrhythmias (and SCD) may also occur in young, otherwise-healthy individuals who are not known to have structural heart disease. In fact, ventricular arrhythmia is the most common cause of sudden death in otherwise-healthy individuals.
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited disorder in individuals with structurally-normal hearts. It is characterized by stress-induced ventricular tachycardia—a lethal arrhythmia that may cause SCD. In subjects with CPVT, physical exertion and/or stress induce bidirectional and/or polymorphic ventricular tachycardias that lead to SCD in the absence of detectable structural heart disease (Laitinen et al., Mutations of the cardiac ryanodine receptor (RyR2) gene in familial polymorphic ventricular tachycardia. Circulation, 103:485-90, 2001; Leenhardt et al., Catecholaminergic polymorphic ventricular tachycardia in children: a 7-year follow-up of 21 patients. Circulation, 91:1512-19, 1995; Priori et al., Clinical and molecular characterization of patients with catecholaminergic polymorphic ventricular tachycardia. Circulation, 106:69-74, 2002; Priori et al., Mutations in the cardiac ryanodine receptor gene (hRyR2) underlie catecholaminergic polymorphic ventricular tachycardia. Circulation, 103:196-200, 2001; Swan et al., Arrhythmic disorder mapped to chromosome 1q42-q43 causes malignant polymorphic ventricular tachycardia in structurally normal hearts. J. Am. Coll. Cardiol., 34:2035-42, 1999).
CPVT is predominantly inherited in an autosomal-dominant fashion. Individuals with CPVT have ventricular arrhythmias when subjected to exercise, but do not develop arrhythmias at rest. Linkage studies and direct sequencing have identified mutations in the human RyR2 gene, on chromosome 1q42-q43, in individuals with CPVT (Laitinen et al., Mutations of the cardiac ryanodine receptor (RyR2) gene in familial polymorphic ventricular tachycardia. Circulation, 103:485-90, 2001; Priori et al., Mutations in the cardiac ryanodine receptor gene (hRyR2) underlie catecholaminergic polymorphic ventricular tachycardia. Circulation, 103:196-200, 2001; Swan et al., Arrhythmic disorder mapped to chromosome 1q42-q43 causes malignant polymorphic ventricular tachycardia in structurally normal hearts. J. Am. Coll. Cardiol., 34:2035-42, 1999).
There are three types of ryanodine receptors, all of which are highly-related Ca2+ channels. RyR1 is found in skeletal muscle, RyR2 is found in the heart, and RyR3 is located in the brain. The type 2 ryanodine receptor (RyR2) is the major Ca2+-release channel required for EC coupling and muscle contraction in cardiac striated muscle.
RyR2 channels are packed into dense arrays in specialized regions of the SR that release intracellular stores of Ca2+, and thereby trigger muscle contraction (Marx et al., Coupled gating between individual skeletal muscle Ca2+ release channels (ryanodine receptors). Science, 281:818-21, 1998). During EC coupling, depolarization of the cardiac-muscle cell membrane, in phase zero of the AP, activates voltage-gated Ca2+ channels. In turn, Ca2+ influx through these channels initiates Ca2+ release from the SR via RyR2, in a process known as Ca2+-induced Ca2+ release (Fabiato, A., Calcium-induced release of calcium from the cardiac sarcoplasmic reticulum. Am. J. Physiol., 245:C1-C14, 1983; Nabauer et al., Regulation of calcium release is gated by calcium current, not gating charge, in cardiac myocytes. Science, 244:800-03, 1989). The RyR2-mediated, Ca2+-induced Ca2+ release then activates the contractile proteins which are responsible for cardiac muscle contraction.
RyR2 is a protein complex comprising four 565,000-dalton RyR2 polypeptides in association with four 12,000-dalton FK506 binding proteins (FKBPs), specifically FKBP12.6 (calstabin). FKBPs are cis-trans peptidyl-prolyl isomerases that are widely expressed, and serve a variety of cellular functions (Marks, A. R., Cellular functions of immunophilins. Physiol. Rev., 76:631-49, 1996). FKBP12 proteins are tightly bound to, and regulate the function of, the skeletal ryanodine receptor, RyR1 (Brillantes et al., Stabilization of calcium release channel (ryanodine receptor) function by FK506-binding protein. Cell, 77:513-23, 1994; Jayaraman et al., FK506 binding protein associated with the calcium release channel (ryanodine receptor). J. Biol. Chem., 267:9474-77, 1992); the cardiac ryanodine receptor, RyR2 (Kaftan et al., Effects of rapamycin on ryanodine receptor/Ca(2+)-release channels from cardiac muscle. Circ. Res., 78:990-97, 1996); a related intracellular Ca2+-release channel, known as the type 1 inositol 1,4,5-triphosphate receptor (IP3R1) (Cameron et al., FKBP12 binds the inositol 1,4,5-trisphosphate receptor at leucine-proline (1400-1401) and anchors calcineurin to this FK506-like domain. J. Biol. Chem., 272:27582-88, 1997); and the type I transforming growth factor β (TGFβ) receptor (TβRI) (Chen et al., Mechanism of TGFbeta receptor inhibition by FKBP12. EMBO J., 16:3866-76, 1997). FKBP12.6 binds to the RyR2 channel (one molecule per RyR2 subunit), stabilizes RyR2-channel function (Brillantes et al., Stabilization of calcium release channel (ryanodine receptor) function by FK506-binding protein. Cell, 77:513-23, 1994), and facilitates coupled gating between neighboring RyR2 channels (Marx et al., Coupled gating between individual skeletal muscle Ca2+ release channels (ryanodine receptors). Science, 281:818-21, 1998), thereby preventing aberrant activation of the channel during the resting phase of the cardiac cycle.
It is clear that leaks in RyR2 channels are associated with a number of pathological states—in both diseased hearts and structurally-normal hearts. Accordingly, methods to repair the leaks in RyR2 could treat or prevent heart failure, cardiac arrhythmias, and sudden cardiac death in millions of patients.
The 1,4-benzothiazepine derivative, JTV-519, or 4-[3-(4-benzylpiperidin-1-yl)propionyl]-7-methoxy-2,3,4,5-tetrahydro-1,4-benzothiazepine monohydrochloride (also known as k201 or ICP-Calstan 100), is a new modulator of calcium-ion channels. In addition to regulating Ca2+ levels in myocardial cells, JTV-519 also modulates the Na+ current and the inward-rectifier K+ current in guinea pig ventricular cells, and inhibits the delayed-rectifier K+ current in guinea pig atrial cells. Studies have shown that JTV-519 has a strong cardioprotective effect against catecholamine-induced myocardial injury, myocardial-injury-induced myofibrillar overcontraction, and ischemia/reperfusion injury. In experimental myofibrillar overcontraction models, JTV-519 demonstrated greater cardioprotective effects than propranolol, verapamil, and diltiazem. Experimental data have also suggested that JTV-519 effectively prevents ventricular ischemia/reperfusion by reducing the level of intracellular Ca2+ overload in animal models.